A quantum-chemical approach to develop tetrahydroquinoxaline as potent ferroptosis inhibitors

Article abstract of DOI:10.1016/j.molstruc.2020.129485

Ferroptosis is a recently characterized form of regulated necrosis with the iron-dependent accumulation of (phospho)lipid hydroperoxides (LOOH). It has attracted considerable attention for its putative involvement in diverse pathophysiological processes, such as cardiovascular disease and neurodegeneration. Here we describe the discovery of tetrahydroquinoxaline, a novel scaffold of ferroptosis inhibitors based on quantum chemistry methods. Tetrahydroquinoxaline deviates showed very good inhibition of ferroptosis, while being non cytotoxic for human cancer cells. And, the advantage of them is their small molecular weight (MW. = 148 Da) that can be coupled with other drugs to form multi-target drugs to better meet the treatment of complicated diseases.

Full text of DOI:10.1016/j.molstruc.2020.129485

Journal of Molecular Structure 1228 (2021) 129485
Contents lists available at ScienceDirect
Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstr
A quantum-chemical approach to develop tetrahydroquinoxaline as
potent ferroptosis inhibitors
Hong-Xu Leia,1, KaiLi Zhangb,1, Yu-Xi Qina, Rong-Jian Dongc, De-Zhan Chena,
HaiFeng Zhoub,∗, Xie-Huang Shenga,∗
a College of Chemistry, Chemical Engineering and Materials Science, Collaborative Innovation Center of Functionalized Probes for Chemical Imaging in
Universities of Shandong, Key Laboratory of Molecular and Nano Probes, Ministry of Education, Shandong Provincial Key Laboratory of Clean Production of
Fine Chemicals, Shandong Normal University, Jinan 250014, PR China
b Hubei Key Laboratory of Natural Products Research and Development, College of Biological and Pharmaceutical Sciences, China Three Gorges University,
Yichang 443002, PR China
c Pingyang center for food and drug control, Wenzhou 325410, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
Ferroptosis is a recently characterized form of regulated necrosis with the iron-dependent accumulation
of (phospho)lipid hydroperoxides (LOOH). It has attracted considerable attention for its putative involve-
ment in diverse pathophysiological processes, such as cardiovascular disease and neurodegeneration. Here
we describe the discovery of tetrahydroquinoxaline, a novel scaffold of ferroptosis inhibitors based on
quantum chemistry methods. Tetrahydroquinoxaline deviates showed very good inhibition of ferroptosis,
while being non cytotoxic for human cancer cells. And, the advantage of them is their small molecular
weight (MW. = 148 Da) that can be coupled with other drugs to form multi-target drugs to better meet
the treatment of complicated diseases.
Received 22 July 2020
Revised 14 October 2020
Accepted 17 October 2020
Available online 28 October 2020
© 2020 Elsevier B.V. All rights reserved.
1. Introduction
ADME properties, and in vivo efficacy by modifying the scaffold of
Fer-1, such as UAMC-2418, UAMC-3023 (Fig 1). [14,15], another ef-
Ferroptosis is a form of regulated non-apoptotic cell death in-
volving overwhelming iron-dependent lipid peroxidation, which is
genetically, biochemically, and morphologically distinct from other
cell death, including apoptosis, unregulated necrosis, necroptosis
and autophagy [1–3]. The process of ferroptosis is essentially de-
stroying the intracellular redox balance associated with the gul-
tathione (GSH) depletion and inactivation of the glutathione per-
oxidase 4 (GPX4) [4–6] and has been implicated in various patho-
logical conditions of the brain, kidney, liver and heart. [7–9] There-
fore, inhibition of the cell death triggered exclusively by ferropto-
sis may represent a promising therapeutic approach for these still
poorly treatable diseases [10,11]. Thus, it is urgent to develop novel
ferroptosis inhibitors.
fort was to test the anti-ferroptotic cell death activity of the exist-
ing compounds in house (the lower line of Fig 1) [16]. However, no
scaffolds of ferroptosis inhibitors were discovered by rational drug
design approaches.
Experimental and computational studies on the mechanism by
which Fer-1 and Lip-1 subvert ferroptosis pointed strongly at their
ability to prevent lipid peroxidation by trapping chain-carrying
peroxyl radicals at endoplasmic reticulum [17–20].
Inhibition of Autoxidation by Radical-Trapping Antioxidants,
kinh
ROO· + AH → ROOH + A·
ROO : lipid peroxidation; AH: radical-trapping antioxidants
And the efficacy as radical-trapping antioxidants (RTAs) relies
heavily on the ability of stability to one-electron oxidation [21,22].
As shown in Fig. 2 and Tab S1, the exceptionally high potency
of Lip-1 and 1,2-dihydroquinoline are probably attributed to the
extensive delocalization of unpaired electrons in radical products
for minimizing the entropic cost of N–H bond breaking [16,17,20].
Much of this resonance stabilization of the unpaired electron is
considered to result from the strong π-π conjugation interactions
in molecule of Lip-1, 1,2-dihydroquinoline (Fig 2). However, com-
pound 1,2,3,4-tetrahydroquinoline does not show any protective ef-
Ferrostatin-1 (Fer-1, Fig 1) and liproxstatin-1 (Lip-1, Fig 1), the
first inhibitors of ferroptosis, were identified by high-throughput
screening methods [12,13]. In the subsequent development of new
inhibitors, one effort was to improve the chemical stability, good
∗
Corresponding authors.
E-mail addresses: zhouhf@ctgu.edu.cn (H. Zhou), shengxiehuang@sdnu.edu.cn
(X.-H. Sheng).
1
These authors contributed equally.
https://doi.org/10.1016/j.molstruc.2020.129485
0022-2860/© 2020 Elsevier B.V. All rights reserved.

H.-X. Lei, K. Zhang, Y.-X. Qin et al.
Journal of Molecular Structure 1228 (2021) 129485
Fig. 1. Chemical structure of the reported ferroptosis inhibitors
Fig. 2. Spin density of the radical products of 1,2,3,4-tetrahydroquinoline, 1,2-dihydro- quinoline and Lip-1. (molecular orbitals are rendered at an isovalue of 0.002)
fect against ferroptosis at all for the reason of no π-π conjugation
interactions in molecule.
As we all known that p-π conjugation and π-π conjugation are
the two main approach to enhance the ability of electron delocal-
ization. But, whether p-π conjugation can be as the predominant
factor in stabilizing the radical product and exhibiting high anti-
ferroptotic cell death activity has not been reported.
In our pursuit to discover novel ferroptosis inhibitors, the scaf-
fold of tetrahydro -quinoxaline was initially identified as power-
ful ferroptosis inhibitors through testing of the compounds with
p-π conjugation. Further optimization of this tetrahydroquinoxa-
line template led to the discovery of a series of novel ferroptosis
inhibitors with high potency. In this report, we will describe in de-
tail the design, syntheses and structure-activity relationship (SAR)
analysis of these novel high potent ferroptosis inhibitors.
Fig. 3. The aminic antioxidants with p-π conjugation.
2. Results and discussion
2.1. Discovery of tetrahydroquinoxaline as ferroptosis inhibitors
peroxyl radical trapping reaction (see Supporting Information for
In order to access the importance of p-π conjugation in sup-
pressing ferroptotic cell death, 6 arylamines are chosen for test-
ing listed in Fig 3. Some are commercially available (1-4), while
the others were synthesized in this work. These compounds are
grouped into three categories: 2H-benzothiazine, 2H-benzoxazine,
and tetrahydroquinoxaline. Their potency for inhibiting erastin-
induced ferroptosis was evaluated in an assay with HT-1080 fi-
brosarcoma cells. In addition to biological evaluations, some quan-
tum chemical parameters, such as reactive energy barrier in lipid
specific details) and E(2) values of p-π conjugation, are also calcu-
lated.
Interestingly, the inhibitory potency of these compounds
against erastin-induced ferroptosis is closely paralleled the E(2)
values, shown in Fig 4 and Tab S1. The p-orbital of nitrogen in
tetrahydroquinoxaline 5 and 6 are most overlap to the aromatic π-
system with the nN→π∗ interaction energies (E(2) values) of 33.60
and 34.79 Kcal•mol−1, respectively. In this case, compound 5 and 6
require a lower energy barrier in the process of trapping lipid per-
2

H.-X. Lei, K. Zhang, Y.-X. Qin et al.
Journal of Molecular Structure 1228 (2021) 129485
Fig. 4. The relationships between anti-ferroptotic cell death activity and the extent of p-π conjugation.
Table 1
Antiferroptotic Activity of Synthesized tetrahydroquinoxalines Library in
Response to Erastin-Induced Ferroptosis in HT-1080 Human Fibrosar-
coma Cells
Comp.
R1
R2
R3
EC50 (nM)
5
-
methyl
methyl
methyl
methyl
methyl
methyl
methyl
methyl
benzyl
ethyl
52
95
42
52
110
91
203
28
85
225
165
37
42
40
17
36
41
36
40
32
45
50
3
2
6
6-Br
methyl
16
17
18
19
20
21
22
37
38
39
40
41
42
43
44
45
46
47
48
Fer-1a
-
-
1
6-Br
-
2
6-Cl
methyl
5
6-F, 7-F
methyl
1
6-ethyl aetate
methyl
15
-
-
2
2
6-Cl
ethyl
6-CN
-
-
-
-
-
-
-
-
-
-
-
-
benzyl
benzyl
benzyl
benzyl
benzyl
benzyl
benzyl
benzyl
23
2
Fig. 5. Cytotoxicity of selected compounds in HepG2 cells.
6-CF3
6-Cl
3
2
1
1
2
3
2
3
2
3
6-F
6-Br
6-butyl
6-F, 7-F
6-Cl, 7-Cl
ꢀ
-
-
-
-
-
benzyl-4 -ethyl
ꢀ
benzyl-4 -Pr
ꢀ
ꢀ
benzyl-4 -Br
benzyl-3 -F
-
oxyl radical and display high inhibitory potency against ferroptosis,
with EC50 values of 52 and 95 nM (Tab S1), respectively, which are
similar with potency of Fer-1. However, the introduction of oxygen
and sulfur of 2H-benzoxazines (3 and 4) and 2H-benzothiazines (1
and 2) significantly decrease the n→π∗ interaction energies and
weaken the ability to stabilize one-electron oxidation of radical
products. Obviously, their inhibitory potency decreased remarkably,
with nearly 5-fold and 50-fold less potent than compound 5, re-
spectively. These results not only elucidate the importance of p-
π conjugation in trapping lipid peroxyl radical, but also identify a
novel scaffold of ferroptosis inhibitors—tetrahydroquinoxaline
The newly developed series of tetrahydroquinoxaline inhibitors
were synthesized by two reduction methods displayed in Scheme
1, 2.
The first method was B2(OH)4-mediated one-pot synthesis of
tetrahydroquinoxalines from 2-aminoanilines and dicarbonyl com-
pounds in water [23] resulted in target compounds 5,6,16-22
(Scheme 1).
The syntheses of the 2-phenyl-tetrahydroquinoxaline contain-
ing compounds 37-48 were accomplished as depicted in scheme
2. 3,4-diaminobenzonitrile 23 effectively reacted with various 2-
bromo-1-phenylethanone analogs in basic conditions to give inter-
mediates 25-36. Cu-catalyzed reduction of 2-phenyl-quinoxalines
with diboronic acid as reductant in an aprotic solvent resulted in
37-48.
2.2. Compounds design and synthesis
In order to verify tetrahydroquinoxaline is a lead scaffold of fer-
roptosis inhibitors and to develop a more potent candidate, two
types of modifications were introduced in this series: (1) replace-
ment of R1, or R3 by electron-donating groups (EDGs) or electron-
withdrawing groups (EWGs), to evaluate the effect of electron den-
sity on the parent structure’s reactivity, (2) addition of alkyl sub-
stituents to the R2 and R3 simultaneously, to evaluate the effect of
steric hindrance on the reactive -NH group.
2.3. Inhibition of erastin-induced ferroptosis
The newly synthesized tetrahydroquinoxaline analogues were
evaluated for prevention of erastin-induced ferroptotic cell death,
and EC50 values are presented in Table 1. In general, all compounds
3

H.-X. Lei, K. Zhang, Y.-X. Qin et al.
Journal of Molecular Structure 1228 (2021) 129485
are found to exhibit potent inhibiting activity against ferroptosis, at
the range of 17-225 nM.
2.4. The cell toxicity of tetrahydroquinoxaline analogues
In order to better describe the structure-activity relationship
of the synthesized tetrahydroquinoxaline analogues, we set com-
pound 16 (EC50 = 42 nM) as the baseline for comparison. As ex-
pected, the introduction of electron-donating groups (EDGs) on
aromatic ring substantially increases the inhibitory potency. Com-
pound 42 possesses greater inhibitory potency against erastin-
induced ferroptosis and greater stability to one-electron oxidation
compared to 16. Compound 42 was identified as the most po-
tent inhibitor of ferroptosis in this work, with EC50 values of 17
nM. Whereas, we observed a loss of potency when substituted by
electron-withdrawing groups (EWGs). Weak EWGs, such as F, Cl,
Br, weaken the inhibitory potency slightly. However, substitution at
the 6-position with CF3, ester or cyanide groups (strong EWGs, 20,
37, 38) yields EC50 = 165 nM, 225 nM and 203 nM, respectively—
roughly one order of magnitude lower potency than compound 42.
For compound 21, 45-48, they showed equivalent or a little better
potency than 16 when introduced EDGs on R3.
Arylamine toxicity is an important concern in the development
of arylamine-containing drugs [24,25]. Thus, in order to comple-
ment the antiferroptotic potential of tetrahydroquinoxalines, we
carried out the studies of detecting the cytotoxicity with HepG2
cells. [26,27] Like Fer-1, the test compounds are cytotoxic only
at concentrations that are significantly (up to 1000-fold) higher
than the concentrations at which they are cytoprotective. The half-
inhibitory concentration (IC50) values of all test compounds are
larger than 150 μM. Compound 21 and 42 display the lowest cy-
totoxicity with no significant diminution in cell viability up to 400
μM. These results indicate that the tetrahydroquinoxalines would
provide an attractive therapeutic window.
3. Conclusions
A novel structural class of ferroptosis inhibitors, tetrahydro-
quinoxaline analogues (compound 5, 6, 16-22, 37-48), are discov-
ered through quantum chemistry and in vitro assay. Besides, as a
novel scaffold of ferroptosis inhibitors, tetrahydroquinoxaline also
possess some other promising features. Firstly, Tetrahydroquinox-
aline analogues have high potency and being low toxicity com-
pounds at the same time, which remarkably reduces the concern
about arylamine toxicity for such structures. Secondly, the tetrahy-
droquinoxaline scaffold is a small structure in which the small-
est active compound (16) has a molecular weight of only 148 Da.
Moreover, this scaffold has multiple modifiable sites that can be
coupled with other drugs to form multi-target drugs to better meet
the treatment of complicated diseases. For example, if the small
compound 16 is coupled with the central acetylcholinesterase in-
hibitors, such as donepezil, rivastigmine, the new molecules would
have a dual-function to treat Alzheimer’s disease. In general, these
novel compounds reported in this study will provide new insights
for the design and development of potent ferroptosis inhibitors.
We further examined the effect of steric hindrance on in-
hibitory potency against ferroptosis. Introduction of alkyl on R2
and R3 simultaneously, such as compound 5, 22, are slightly less
potent than only one substitution on R3 (16). In order to check the
steric effect of benzene ring, quantum chemistry was employed to
analysis the site selectivity for compound 21. As shown in Fig S1,
the functional –NH group closed to the benzene ring showed ap-
parently unfavorable than the other –NH group. It can be stated
that benzene ring substitution produce a steric hindrance effect on
the reactivity of adjacent -NH group. Therefore, just single sub-
stitution at positions R2 or R3 would be a more recommended
choice.
In conclusion, consistent with the reactivity of other aminic
RTAs, tetrahydroquinoxaline substituted with EDGs increase the
potency and EWGs decrease the potency. The inhibitory potency
for tetrahydroquinoxaline will be partially blocked by the steric
hindrance effect of simultaneous substitution on positions R2 and
R3.
4

H.-X. Lei, K. Zhang, Y.-X. Qin et al.
4. Methods
Journal of Molecular Structure 1228 (2021) 129485
Methyl-1,2,3,4-tetrahydroquinoxaline (16). By following general
procedure A and using 2-aminoaniline (108 mg, 1 mmol), 2-
oxopropanal (72 mg, 1 mmol) as the staring material, the forma-
tion of the 2-Methyl-1,2,3,4-tetrahydroquinoxaline 16 (124 mg, 1
mmol) was achieved (yield 84%); yellow solid. m.p. 72-74°C; 1H
NMR (400 MHz, CDCl3) δ= 6.67-6.64 (m, 2H), 6.58-6.55 (m, 2H),
3.58-3.34 (m, 4H), 3.08 (dd, J = 10.8, 8.4 Hz, 1H), 1.23 (d, J = 6.4
Hz, 3H) ppm; 13C NMR (100 MHz, CDCl3) δ= 133.62, 133.25, 118.71,
114.53, 114.48, 48.28, 45.74, 19.96 ppm.
4.1. Reagents and compounds synthesis
Fer-1 and Erastin were obtained from Sigma-Aldrich (Sigma
Chemicals St. Louis, MO, USA). Compound 1-4 were purchased
from Alfa Aesar (Ward Hill, MA, USA). All reagents were obtained
from commercial sources and were used without further purifi-
cation unless otherwise stated. Acetone was dried with Type 4A
Linde molecular sieves and then distilled. All reactions were mon-
itored by thin-layer chromatography (TLC). 1H NMR and 13C NMR
spectra were measured on a Bruker 400 MHz spectrometer, using
deuterated dimethyl sulfoxide (DMSO-d6) as solvent and TMS as
internal standard.
Bromo-2-methyl-1,2,3,4-tetrahydroquinoline (17): By following
general procedure A and using 4-bromobenzene-1,2-diamine (186
mg,
1 mmol), 2-oxopropanal (72 mg, 1 mmol) as the star-
ing material, the formation of the 6-bromo-2-methyl-1,2,3,4-
tetrahydroquinoline 17 (197 mg, 1 mmol) was achieved (yield
87%); Colorless oil. 1H NMR (400 MHz, CDCl3): δ = 7.05-7.10 (m,
2H), 6.38 (d, J = 8.4 Hz, 1H), 3.39-3.44 (m, 1H), 2.74-3.00 (m, 2H),
1.93-1.98 (m, 1H), 1.57-1.62 (m, 1H), 1.24 (d, J = 6.4 Hz, 3H) ppm;
13C NMR (100 MHz, CDCl3): δ = 143.79, 131.69, 129.34, 123.15,
115.40, 108.29, 47.11, 29.64, 26.43, 22.49 ppm.
The following sections comprise the synthetic procedures and
analytical data for all compounds reported in this manuscript. Sev-
eral synthesis procedures that were used in the preparation of in-
termediates and final products are summarized here as “General
Procedures”.
General Procedure A: A flask was charged with 2-aminoaniline,
2,3-butanedione, B2(OH)4 (8 mmol, 720 mg, 8 eq.) and water (3
mL) under N2. The reaction was stirred at 80°C for 4 h. When the
reaction was complete monitored by TLC, the mixture was cooled
to room temperature, extracted with ethyl acetate (3 × 20 mL). The
combined organic phase was washed with water, dried over anhy-
drous Na2SO4, filtered, and concentrated under reduced pressure
to give a crude product. After determination of the diastereomeric
excess by 1H NMR, the crude product was purified by silica gel
column chromatography to give the product 5,6,16-22.
Chloro-2,3-dimethyl-1,2,3,4-tetrahydroquinoxaline (18). By follow-
ing general procedure A and using 4-chlorobenzene-1,2-diamine
(142 mg, 1 mmol), 2,3-butanedione (86 mg, 1 mmol) as the star-
ing material, the formation of the 6-Chloro-2,3-dimethyl-1,2,3,4-
tetrahydroquinoxaline 18 (141 mg, 1 mmol) was achieved (yield
72%); White solid. m.p.146-147°C; 1H NMR (400 MHz, CDCl3)
δ = 6.54 (dd, J = 8.4, 2.2 Hz, 1H), 6.49 (d, J = 2.2 Hz, 1H), 6.43
(d, J = 8.4 Hz, 1H), 3.05-2.99 (m, 2H), 1.19 (dd, J = 6.0, 1.2 Hz, 6H)
ppm; 13C NMR (100 MHz, CDCl3) δ: 134.51, 131.85, 123.03, 117.79,
114.46, 113.26, 51.90, 18.95, 18.92 ppm.
General Procedure B: A flask was charged with benzene-1,2-
diamine, 2-bromo-1- phenylethanone, NaHCO3 (1.2 mmol, 220
mg.) and DMSO (5 mL). The reaction was stirred at 120°C for 24
h. The mixture was cooled to room temperature and quenched
with water, extracted with ethyl acetate (3 × 20 mL). The com-
bined organic phase was washed with water, dried over anhydrous
Na2SO4, filtered, and concentrated under reduced pressure to give
a crude product. The crude product was purified by silica gel col-
umn chromatography to give the intermediate 25-36. A 20 mL
Schlenk tube was charged with 2-phenylquinoxaline, Cu(OAc)2 (4.5
mg, 0.025 mmol), B2(OH)4 (135 mg, 3 mmol), and MeCN (3.0 mL).
The mixture was stirred at 80 °C for 12 h. When the reaction
was complete monitored by TLC, the mixture was cooled to room
temperature, extracted with ethyl acetate (3 × 20 mL). The com-
bined organic phase was washed with water, dried over anhydrous
Na2SO4, filtered, and concentrated under reduced pressure to give
a crude product. The crude product was purified by silica gel col-
umn chromatography to give the product 37-48.
6,7-Difluoro-2,3-dimethyl-1,2,3,4-tetrahydroquinoxaline (19): By
following general procedure A and using 4,5-difluorobenzene-1,2-
diamine (144 mg, 1 mmol), 2,3-butanedione (86 mg, 1 mmol)
as the staring material, the formation of the 6,7-Difluoro-2,3-
dimethyl-1,2,3,4-tetrahydroquinoxaline 19 (103 mg, 1 mmol) was
achieved (yield 52%); White solid. m.p.91-92°C; 1H NMR (400
MHz, CDCl3) δ= 6.46 (dd, J = 8.4, 5.2 Hz, 1H), 6.30-6.24 (m, 1H),
3.54-3.47 (m, 2H), 1.16 (d, J = 6.4 Hz, 6H) ppm; 13C NMR (100
MHz, CDCl3) δ= 157.28 (d, J = 233 Hz), 133.90 (d, J = 10 Hz),
127.76, 115.11 (d, J = 9.2 Hz), 103.86 (d, J = 22.3 Hz), 101.01 (d,
J = 25.9 Hz), 49.03, 48.87, 17.07, 16.86 ppm.
Ethyl
2,3-dimethyl-1,2,3,4-tetrahydroquinoxaline-6-carboxylate
and using ethyl 3,4-
(20). By following general procedure
A
diaminobenzoate (180 mg, 1 mmol), 2,3-butanedione (86 mg, 1
mmol) as the staring material, the formation of the Ethyl 2,3-
dimethyl-1,2,3,4-tetrahydroquinoxaline-6-carboxylate 20 (150 mg,
1 mmol) was achieved (yield 64%); White solid. m.p.75-76 °C;
mixture of cis and trans isomer: 1H NMR (400 MHz, CDCl3) δ=
7.40 (dd, J = 8.4, 1.8 Hz, 1H), 7.30 (d, J =1.8 Hz, 1H), 6.46 (dd,
J = 8.4, 4.2 Hz, 1H), 4.33 (dd, J = 14.2, 7.2 Hz, 2H), 3.63-3.48 (m,
2H, cis), 3.16-2.99 (m, 1H, trans), 1.38 (dd, J = 6.4, 8.0 Hz, 3H),
1.22 (dd, J = 6.4, 4.0 Hz, trans), 1.17 (dd, J = 6.4, 4.0 Hz, 4H, cis)
ppm; 13C NMR (100 MHz, CDCl3) δ= 167.09, 138.05 (trans), 137.44
(cis), 132.21 (trans), 131.43 (cis), 121.61 (trans), 121.53 (cis), 119.49
(trans), 119.45 (cis), 115.19 (trans), 114.76 (cis), 112.42 (trans), 112.02
(cis), 60.14, 52.24 (trans), 51.40 (trans), 49.22 (cis), 48.49 (cis), 19.06
(trans), 18.89 (trans), 17.35 (cis), 17.09 (cis), 14.49 ppm.
2,3-Dimethyl-1,2,3,4-tetrahydroquinoxaline (5). By following gen-
eral procedure A and using 2-aminoaniline (108 mg, 1 mmol), 2,3-
butanedione (86 mg, 1 mmol) as the staring material, the forma-
tion of the 2,3-Dimethyl-1,2,3,4-tetrahydroquinoxaline 5 (104 mg,
1 mmol) was achieved (yield 64%); White solid. m.p. 112-114 °C;
1H NMR (400 MHz, CDCl3) δ= 6.62 (dd, J= 5.8, 3.4 Hz, 2H), 6.53
(dd, J = 5.8, 3.4 Hz, 2H), 3.54-3.52 (m, 2H), 1.170 (d, J = 5.6 Hz,
6H); 13C NMR (100 MHz, CDCl3) δ = 132.66, 118.56, 114.42, 49.04,
17.28 ppm.
Bromo-2,3-dimethyl-1,2,3,4-tetrahydroquinoxaline (6). By follow-
ing general procedure A and using 4-bromobenzene-1,2-diamine
(186 mg, 1 mmol), 2,3-butanedione (86 mg, 1 mmol) as the star-
ing material, the formation of the 6-Bromo-2,3-dimethyl-1,2,3,4-
tetrahydroquinoxaline 6 (209 mg, 1 mmol) was achieved (yield
87%); White solid. m.p. 109-111°C; 1H NMR (400 MHz, CDCl3) δ=
6.68 (dd, J = 8.4, 2.2 Hz, 1H), 6.63 (d, J = 2.2 Hz, 1H), 6.39 (d,
J = 8.4 Hz. 1H) 3.07-3.00 (m, 2H), 1.21 (d, J = 4.0 Hz, 6H) ppm; 13C
NMR (100 MHz, CDCl3) δ: 134.85, 132.35, 120.72, 116.00, 114.84,
110.03, 51.84, 51.79, 29.70, 18.94 ppm.
2-Phenyl-1,2,3,4-tetrahydroquinoxaline (21). By following general
procedure A and using 2-aminoaniline (108 mg, 1 mmol), 2-oxo-2-
phenylacetaldehyde (134 mg, 1 mmol) as the staring material, the
formation of the 2-Phenyl-1,2,3,4-tetrahydroquinoxaline 21 (164
mg, 1 mmol) was achieved (yield 78%); Yellow solid. m.p .74-76
°C; 1H NMR (400 MHz, CDCl3) δ= 7.45-7.34 (m, 5H), 6.70-6.67 (m,
2H), 6.65-6.61 (m, 2H), 4.53 (dd, J = 8.0 Hz, 2.8 H, 1H), 3.39 (s,
1H), 3.50 (dd, J = 11.0 Hz, 3.0 Hz, 1H), 3.37 (dd, J = 11.0, 8.4 H,
1H) ppm; 13C NMR (100 MHz, CDCl3) δ= 141.84, 134.18, 132.70,
5

H.-X. Lei, K. Zhang, Y.-X. Qin et al.
Journal of Molecular Structure 1228 (2021) 129485
128.67, 127.94, 127.92, 127.03, 127.00,119.03, 118.81, 114.81, 114.48,
54.76, 49.16 ppm.
NMR (400 MHz, CDCl3) δ=7.50-7.32 (m, 5H), 6.80-6.66 (m, 2H),
6.67-6.59 (m, 1H), 4.57-4.48 (m, 1H), 3.97 (s, 2H), 3.53-3.49 (m,
1H), 3.40-3.30 (m, 1H) ppm. 13C NMR (100 MHz, CDCl3) δ= 142.25,
141.92, 133.71, 130.37, 128.62, 127.85, 127.03, 115.72, 114.72, 111.86,
55.05, 49.45 ppm.
6-chloro-2,3-diethyl-1,2,3,4-tetrahydroquinoxaline (22): By follow-
ing general procedure A and using 4-chlorobenzene-1,2-diamine
(144 mg, 1 mmol), hexane-3,4-dione (114 mg, 1 mmol) as the star-
ing material, the formation of the 6-Chloro-2,3-dimethyl-1,2,3,4-
tetrahydroquinoxaline 22 (161 mg, 1 mmol) was achieved (yield
72%); White solid. m.p.105-107 °C; 1H NMR (400 MHz, DMSO) δ=
6.59-6.14 (m, 3H), 5.70 (s, 1H), 5.49 (s, 1H), 3.08 (dd, J = 5.0, 2.4
Hz, 2H), 1.35-1.28 (m, 4H), 0.93 (td, J = 7.6, 6.4, 3.8 Hz, 6H) ppm.
13C NMR (100 MHz, DMSO) δ = 135.23, 132.62, 120.29, 115.88,
113.95, 112.21, 53.70, 23.13, 10.81 ppm.
(tert-butyl)-2-phenyl-1,2,3,4-tetrahydroquinoxaline
(42):
By
following general procedure A and using 6-(tert-butyl)-2-
phenylquinoxaline (131 mg, 0.5 mmol) as the staring ma-
terial, the formation of the 6-(tert-butyl)-2-phenyl-1,2,3,4-
tetrahydroquinoxaline 42 (71 mg, 0.5 mmol) was achieved (yield
53%); Yellow solid. m.p. 95-97 °C; 1H NMR (400 MHz, CDCl3)
δ=7.45-7.35 (m, 5H), 6.76-6.55 (m, 3H), 4.52 (d, J = 8.6 Hz, 1H),
3.90 (s, 2H), 3.56-3.43 (m, 1H), 3.38-3.35 (m, 1H), 1.32 (s, 9H)
ppm. 13C NMR (100 MHz, CDCl3) δ= 142.25, 141.92, 133.71, 130.37,
128.62, 127.85, 127.03, 115.72, 114.72, 111.86, 55.05, 49.45, 33.98,
31.57 ppm.
Phenyl-1,2,3,4-tetrahydroquinoxaline-6-carbonitrile (37): By fol-
lowing general procedure
B and using 2-phenylquinoxaline-6-
carbonitrile (116 mg, 0.5 mmol) as the staring material, the forma-
tion of the 2-phenyl-1,2,3,4-tetrahydroquinoxaline-6-carbonitrile
37 (108 mg, 0.5 mmol) was achieved (yield 92%); Yellow solid.
m.p.142-144 °C; 1H NMR (400 MHz, DMSO) δ= 7.41-7.28 (m, 5H),
6.86 (d, J = 2.0 Hz, 1H), 6.71 (d, J = 2.0 Hz, 1H), 6.60 (d, J = 8.0 Hz,
1H), 5.98 (s, 1H), 4.52-4.42 (m, 1H), 3.38 (dd, J = 10.0, 2.0 Hz, 2H),
3.07 (dd, J = 12.0, 8.0 Hz, 1H) ppm. 13C NMR (100 MHz, DMSO) δ=
142.54, 139.50, 133.83, 128.78, 127.88, 127.31, 122.71, 121.54, 114.95,
112.81, 97.08, 53.36, 47.07 ppm.
6,7-difluoro-2-phenyl-1,2,3,4-tetrahydroquinoxaline
(43):
By
following general procedure A and using 6,7-difluoro-2-
phenylquinoxaline (121 mg, 0.5 mmol) as the staring ma-
terial, the formation of the 6,7-difluoro-2-phenyl-1,2,3,4-
tetrahydroquinoxaline 43 (100 mg, 0.5 mmol) was achieved
(yield 81%); Yellow solid. m.p. 116-118 °C; 1H NMR (400 MHz,
DMSO) δ= 7.37 (d, J = 4.4 Hz, 4H), 7.34-7.26 (m, 1H), 6.49 (dd,
J = 12.0, 8.0 Hz, 1H), 6.40 (dd, J = 12.0, 8.0 Hz, 1H), 5.96 (s,
1H), 5.72 (s, 1H), 4.28 (dd, J = 8.0, 4.0 Hz, 1H), 3.43-3.27 (m,
2H), 3.06 (dd, J = 11.1, 8.0 Hz, 1H) ppm. 13C NMR (100 MHz,
DMSO) δ=142.73, 131.12 (d, J = 7.2 Hz), 130.26 (d, J = 7.3 Hz),
128.74,127.81, 127.40, 101.42 (dd, J = 20.7, 20.7 Hz), 53.24, 48 ppm.
2-phenyl-6-(trifluoromethyl)-1,2,3,4-tetrahydroquinoxaline(38):
By following general procedure
B
and using 2-phenyl-6-
(trifluoromethyl)quinoxaline (137 mg, 0.5 mmol) as the staring
material, the formation of the 2-phenyl-6-(trifluoromethyl)-1,2,3,4-
tetrahydroquinoxaline 38 (121 mg, 0.5 mmol) was achieved (yield
87%); Yellow solid . m.p. 115-117 °C; 1H NMR (400 MHz, DMSO)
δ= 7.37 (d, J = 4.4 Hz, 4H), 7.32-7,28 (m, 1H), 6.54 (d, J = 2.0 Hz,
1H), 6.45-6.34 (m, 2H), 6.10 (s, 1H), 5.71 (s, 1H), 4.35-4.32 (m, 1H),
3.32 (s, 1H), 3.09-3.04 (m, 1H) ppm. 13C NMR (100 MHz, DMSO)
δ= 142.95, 136.17, 132.81, 128.73, 127.74, 127.33, 120.62, 116.48,
114.26, 112.78, 53.32, 48.07 ppm.
6,7-dichloro-2-phenyl-1,2,3,4-tetrahydroquinoxaline
(44):
By
following general procedure A and using 6,7-dichloro-2-
phenylquinoxaline (137 mg, 0.5 mmol) as the staring ma-
terial, the formation of the 6,7-difluoro-2-phenyl-1,2,3,4-
tetrahydroquinoxaline 44 (117 mg, 0.5 mmol) was achieved
(yield 84%); White solid. m.p. 116-118 °C; 1H NMR (400 MHz,
DMSO) δ= 7.40-7.35 (m, 4H), 7.35-7.27 (m, 1H), 6.67 (s, 1H), 6.58
(s, 1H), 6.27 (s, 1H), 6.09-5.97 (m, 1H), 4.33 (dt, J = 6.0, 1.8 Hz,
1H), 3.35 (s, 1H), 3.11-3.06 (m, 1H) ppm. 13C NMR (101 MHz,
DMSO) δ= 142.54, 135.12, 134.37, 128.78, 127.88, 127.38, 117.73,
117.40, 113.62, 113.24, 52.94, 47.55 ppm.
6-chloro-2-phenyl-1,2,3,4-tetrahydroquinoxaline (39) By following
general procedure B and using 6-chloro-2-phenylquinoxaline (120
mg, 0.5 mmol) as the staring material, the formation of the
6-chloro-2-phenyl-1,2,3,4-tetrahydroquinoxaline 39 (121 mg, 0.5
mmol) was achieved (yield 89%); Yellow solid. m.p.116-117 °C; mix-
ture of cis and trans isomer: 1H NMR (400 MHz, DMSO) δ= 7.37
(d, J = 5.6 Hz, 4H), 7.33-7.26 (m, 1H), 6.66 (s, 0.4 H, trans), 6.57-
6.48 (m, 1H, cis), 6.45-6.35 (m, 2H, cis), 6.10 (s, 0.4 H, trans), 5.72
(s, 1H, cis), 5.49 (s, 0.4 H, trans), 4.32 (s, 1H), 3.33 (d, J = 10.8
Hz, 2H), 3.07 (q, J = 9.0 Hz, 1H) ppm.13C NMR (100 MHz, DMSO)
δ= 142.91, 134.81 (trans), 133.91 (cis), 133.22 (cis), 128.75 (trans),
128.67 (cis), 127.77 (trans), 127.62 (cis), 127.39 (cis), 127.35 (trans),
119.37, 117.60 (trans), 117.38 (cis), 115.43 (trans), 114.75 (cis), 114.06
(trans), 113.71 (cis), 53.67 (cis), 53.22 (trans), 48.63 (cis), 48.01
(trans) ppm.
2-(4-ethylphenyl)-1,2,3,4-tetrahydroquinoxaline (45): By following
general procedure A and using 2-(4-ethylphenyl)quinoxaline (117
mg, 0.5 mmol) as the staring material, the formation of the 2-(4-
ethylphenyl)-1,2,3,4-tetrahydroquinoxaline 45 (107 mg, 0.5 mmol)
was achieved (yield 90%); Yellow solid .m.p. 97-99 °C; 1H NMR
(400 MHz, CDCl3) δ= 7.36 (d, J = 8.0 Hz, 2H), 7.26 (d, J = 8.0
Hz, 2H), 6.72-6.65 (m, 2H), 6.65-6.57 (m, 2H), 4.51 (dd, J = 8.2,
3.0 Hz, 1H), 3.50 (dd, J = 11.0, 3.0 Hz, 1H), 3.37 (dd, J = 11.0,
8.2 Hz, 1H), 2.71 (q, J = 7.6 Hz, 2H), 1.30 (s, 3H) ppm. 13C NMR
(100 MHz, CDCl3) δ= 144.03, 139.07, 134.24, 132.81, 128.16, 127.00,
118.92, 118.74, 114.72, 114.42, 54.49, 49.21, 28.59, 15.67 ppm.
2-(4-propylphenyl)-1,2,3,4-tetrahydroquinoxaline (46): By follow-
ing general procedure A and using 2-(4-propylphenyl)quinoxaline
(124 mg, 0.5 mmol) as the staring material, the formation of
the 2-(4-propylphenyl)-1,2,3,4-tetrahydroquinoxaline 46 (99 mg,
0.5 mmol) was achieved (yield 79%); Yellow solid. m.p. 103-104
°C; 1H NMR (400 MHz, CDCl3) δ= 7.34 (d, J = 8.0 Hz, 2H), 7.22 (d,
J = 8.0 Hz, 2H), 6.72-6.56 (m, 4H), 4.50 (dd, J = 8.2, 3.0 Hz, 1H),
3.55-3.45 (m, 1H), 3.37 (dd, J = 11.0, 8.2 Hz, 1H), 2.63 (dd, J = 8.6,
6.7 Hz, 2H), 1.72-1.65 (m, 2H), 1.30 (s, 2H), 0.99 (t, J = 7.2 Hz, 3H)
ppm. 13C NMR (100 MHz, CDCl3) δ= 142.49, 139.03, 134.27, 132.72,
128.74, 126.89, 118.96, 118.73, 114.76, 114.42, 54.49, 49.19, 37.75,
24.61, 13.91 ppm.
6-fluoro-2-phenyl-1,2,3,4-tetrahydroquinoxaline (40): By follow-
ing general procedure A and using 6-fluoro-2-phenylquinoxaline
(112 mg, 0.5 mmol) as the staring material, the formation of
the 6-fluoro-2-phenyl-1,2,3,4-tetrahydroquinoxaline 40 (87 mg, 0.5
mmol) was achieved (yield 76%); Yellow solid. m.p.109-111 °C; 1H
NMR (400 MHz, DMSO) δ= 7.38 (d, J = 4.0, 4H), 7.30 (dd, J = 8.0,
4.0 Hz, 1H), 6.45-6.30 (m, 2H), 6.23-6.14 (m, 1H), 6.12 (s, 1H), 5.42
(s, 1H), 4.36 (dd, J = 7.6, 3.2 Hz, 1H), 3.31 (dd, J = 11.0, 3.2 Hz, 1H),
3.04 (dd, J = 11.0, 7.6 Hz, 1H) ppm. 13C NMR (100 MHz, DMSO)
δ= 155.96 (d, J = 229 Hz), 143.08, 136.13 (d, J = 10.4 Hz), 130.03,
128.70, 127.71, 127.26, 113.78 (d, J = 9.0 Hz), 102.29 (d, J = 22.3
Hz), 100.09 (d, J = 25.1 Hz), 53.68, 48.33 ppm.
6-bromo-2-phenyl-1,2,3,4-tetrahydroquinoxaline (41): By follow-
ing general procedure A and using 6-bromo-2-phenylquinoxaline
(142 mg, 0.5 mmol) as the staring material, the formation of the
6-bromo-2-phenyl-1,2,3,4-tetrahydroquinoxaline 41 (124 mg, 0.5
mmol) was achieved (yield 86%); Yellow solid. m.p.102-104 °C; 1H
bromophenyl)-1,2,3,4-tetrahydroquinoxaline (47): By following
general procedure A and using 2-(4-bromophenyl)quinoxaline (142
mg, 0.5 mmol) as the staring material, the formation of the 2-(4-
bromophenyl)-1,2,3,4-tetrahydroquinoxaline 47 (108 mg, 0.5 mmol)
6

H.-X. Lei, K. Zhang, Y.-X. Qin et al.
Journal of Molecular Structure 1228 (2021) 129485
was achieved (yield 75%); Yellow solid. m.p.145-147 °C; 1H NMR
(400 MHz, CDCl3) δ = 7.58-7.49 (m, 2H), 7.31 (d, J = 8.4 Hz, 2H),
6.73-6.67 (m, 2H), 6.67-6.59 (m, 2H), 4.50 (dd, J = 8.0, 3.0 Hz,
1H), 3.48 (dd, J = 11.0, 3.0 Hz, 1H), 3.32 (dd, J = 11.0, 8.0 Hz, 1H)
ppm. 13C NMR (100 MHz, CDCl3) δ= 140.98, 133.77, 132.71, 131.77,
128.72, 121.70, 119.08, 119.01, 114.81, 114.55, 54.17, 48.95 ppm.
(3-fluorophenyl)-1,2,3,4-tetrahydroquinoxaline (48): By following
general procedure A and using 2-(4-fluorophenyl)quinoxaline (112
mg, 0.5 mmol) as the staring material, the formation of the 2-(3-
fluorophenyl)-1,2,3,4-tetrahydroquinoxaline 48 (88 mg, 0.5 mmol)
was achieved (yield 77%); Yellow solid. m.p.96-98 °C; 1H NMR (400
MHz, CDCl3) δ =7.37 (td, J = 8.0, 5.8 Hz, 1H), 7.25-7.12 (m, 2H),
7.09-6.99 (m, 1H), 6.75-6.67 (m, 2H), 6.67-6.58 (m, 2H), 4.54 (dd,
J = 8.0, 3.2 Hz, 1H), 3.56-3.47 (m, 1H), 3.37-3.32 (m, 1H) ppm. 13C
NMR (100 MHz, CDCl3) δ= 164.35, 161.91, 144.69, 133.74, 132.72,
130.14, 130.06, 122.58, 122.55, 119.10, 119.02, 114.83, 114.63, 114.57,
114.00, 113.78, 54.32, 48.95 ppm.
Software, Data curation. HaiFeng Zhou: Funding acquisition, Re-
sources. Xie-Huang Sheng: Supervision, Writing - review & edit-
ing.
Acknowledgments
This work is supported by the National Natural Science Foun-
dation of China (NO. 11774207, 11574184), the Primary Research &
Development Plan of Shandong Province (NO. 2015GSF118175) and
National Undergraduates’ Innovation and Entrepreneurship Train-
ing Program (No. S202010445042) .
Supplementary materials
Supplementary material associated with this article can be
found, in the online version, at doi:10.1016/j.molstruc.2020.129485.
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